Electron microscopy of cervical-mucus glycoproteins and fragments therefrom. The use of colloidal gold to make visible 'naked' protein regions.
Subunits of human cervical-mucus glycoproteins obtained by reductive cleavage of whole mucins and high-Mr glycopeptides (T-domains) obtained after their trypsin digestion were studied with electron microscopy after spreading the macromolecules in a monolayer of benzyldimethylalkylammonium chloride. Subunits were observed as linear and apparently flexible particles, with number- and weight-average lengths of 390 nm and 460 nm respectively. T-domains randomly distributed on the grid have number- and weight-average lengths of 90 nm and 103 nm respectively, whereas when aligned (possibly stretched by flow) they were longer, with number-average and weight-average lengths of 150 nm and 170 nm respectively. Subunits complexed with gold appeared as segmented structures, with a distribution of inter-gold distances similar to the length distribution for the relaxed T-domains. The whole mucins had few binding sites for gold, suggesting that reduction exposes hydrophobic protein-rich regions with high affinity for gold. Most T-domains had a binding site at one end, indicating the presence of a residual protruding naked peptide region. We conclude that mucins are assembled from subunits joined end-to-end, and that each subunit consists of alternating oligosaccharide 'clusters' (approx. 100 nm) and naked peptide regions which have (after reduction) a high affinity for colloidal gold.
Sheehan
J K
JK
Department of Biochemistry and Molecular Biology, University of Manchester, U.K.
Carlstedt
I
I
D002582:Cervix Mucus / Q000378:metabolism*; D005260:Female; D006023:Glycoproteins / Q000648:ultrastructure*; D006046:Gold; D006801:Humans; D007150:Immunohistochemistry
D006023:Glycoproteins; D006046:Gold